How cis-Acyltransferase Assembly-Line Ketosynthases Gatekeep for Processed Polyketide Intermediates
How cis-Acyltransferase Assembly-Line Ketosynthases Gatekeep for Processed Polyketide Intermediates
复制标题
DOI：
10.1021/acschembio.1c00598
复制
发表时间：
2021-09-30
影响因子：
4
通讯作者：
Keatinge-Clay, Adrian T.
中科院分区：
文献类型：
--
作者：
Hirsch, Melissa;Fitzgerald, Brendan J.;Keatinge-Clay, Adrian T.
随着聚酮合成酶(PKS)模块的重新定义，对其最下游结构域--酮合成酶(KS)的新认识正在出现。除了发挥其在酰基辅酶A构建块和不断增长的聚酮之间产生碳-碳键的既定作用外，它还可能阻止未完全加工的中间体。在这里，我们调查了来自92个主要放线菌，顺式酰基转移酶装配线的739个KSS。当KSS根据其聚酮底物的α-碳和β-碳的化学成分被分为16个家族时，32个底物隧道残基的比较显示了独特的序列指纹。令人惊讶的是，当考虑到伽马碳的化学作用时，还检测到了额外的指纹。有代表性的K_s被模拟与其天然聚酮底物结合，以更好地理解观察到的模式，例如用较小的残基取代色氨酸以容纳L-α-甲基基团，或用较大的残基取代四个较小的残基以更好地与引物单元或二酮基团接触。在模型三酮合成酶中对KS中保守的谷氨酰胺进行突变表明底物隧道对改变敏感，而设计该KS以接受非自然底物可能需要几次突变。
With the redefinition of polyketide synthase (PKS) modules, a new appreciation of their most downstream domain, the ketosynthase (KS), is emerging. In addition to performing its well-established role of generating a carbon-carbon bond between an acyl-CoA building block and a growing polyketide, it may gatekeep against incompletely processed intermediates. Here, we investigate 739 KSs from 92 primarily actinomycete, cis-acyltransferase assembly lines. When KSs were separated into 16 families based on the chemistries at the alpha- and beta-carbons of their polyketide substrates, a comparison of 32 substrate tunnel residues revealed unique sequence fingerprints. Surprisingly, additional fingerprints were detected when the chemistry at the gamma-carbon was considered. Representative KSs were modeled bound to their natural polyketide substrates to better understand observed patterns, such as the substitution of a tryptophan by a smaller residue to accommodate an L-alpha-methyl group or the substitution of four smaller residues by larger ones to make better contact with a primer unit or diketide. Mutagenesis of a conserved glutamine in a KS within a model triketide synthase indicates that the substrate tunnel is sensitive to alteration and that engineering this KS to accept unnatural substrates may require several mutations.